Field of the Invention
The present invention provides methods for purifying an active polypeptide or immunoconjugate from a solution containing the polypeptide or immunoconjugate and an acidic variant thereof, wherein said acidic variant is a deamidated species of said polypeptide or immunoconjugate. The present invention also provides formulations containing such purified polypeptides or immunoconjugates.
Background Art
The large-scale, economic purification of proteins is a factor for the biopharmaceutical industry. Therapeutic proteins are typically produced using prokaryotic or eukaryotic cell lines that are engineered to express the protein of interest from a recombinant plasmid containing the gene encoding the protein. Separation of the desired protein from the mixture of components fed to the cells and cellular by-products to an adequate purity, e.g., sufficient for use as a human therapeutic, poses a formidable challenge to biologics manufacturers for several reasons.
Manufacturers of protein-based pharmaceutical products must comply with strict regulatory standards, including extremely stringent purity requirements. To ensure safety, regulatory agencies, such as Food and Drug Administration (FDA), require that protein-based pharmaceutical products are substantially free from impurities, including both product related contaminants such as aggregates, fragments and variants of the recombinant protein and process related contaminants such as host cell proteins, media components, viruses, DNA and endotoxins. While various protein purification schemes are available and widely used in the biopharmaceutical industry, they typically include an affinity-purification step, such as Protein A purification in the case of antibodies, in order to reach a pharmaceutically acceptable degree of purity.
The development of a purification scheme applicable to a particular biomolecule or various biomolecules that is scaleable, controllable, and that strategically employs the use of particular resins or a combination of resins will allow its integration into product development at a very early stage in overall drug development. This approach to the design of a purification scheme can minimize costly changes to manufacturing processes which may otherwise be necessary later in drug development or, worse, after approval. As the process is scaled-up and approaches good manufacturing practices (GMP) production conditions, additional inherent complexities arise, including those associated with resin packing and buffer preparation. The manufacturing process, and its capacity, can be improved by simplifying the purification scheme, by eliminating process steps and maximizing throughput and productivity, while maintaining the integrity and purity of the molecule that is being purified. Therefore, it would be desirable and advantageous to start with a simple and efficient process that can produce a drug substance of high quality and safety.
One complexity associated with the purification of a drug product is the maintenance of potency throughout the purification process. Many factors can contribute to a reduction or inhibition of potency, including the modification of the drug product during the development process. Such modification can occur at various stages of the process, for example, when the protein is being expressed in the cell, or when a protein that has been isolated from a cell is subject to various conditions or buffers. The present invention provides a method for purifying an active polypeptide or immunoconjugate from a solution containing a modified variant of the polypeptide or immunoconjugate, where the presence of this modified variant results in an inhibition in potency of the final drug product.